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Javanese Tree Frog (R. margaritifer 1837) is an endemic frog of Java Island. Population of this species is threatened due to human activity and the pathogenic fungus Batrachochytrium dendrobatidis (Bd). Therefor sterilization of Bd is needed as the first effort to protect of R. margaritifer outside their natural habitats. The research aims to assess: 1) frog behaviour after and before sterilization treatment; 2) best temperature for sterilization by heat treatment for Bd on frogs vitality; and 3) behaviour of frog’s adaptation in terrarium after sterilization. Research were conducted in Indonesian Safari Park I Cisarua, West Java includes specimen’s collection in Curug Jaksa and Rumah Dua. Sterilization were carried out using two methods, i.e (I) aquarium heater with hot steam and (II) aquarium heater with dry heat temperature. A total of 12 frogs were sterilized with 3 different temperature treatment for each heater. Ad libitum sampling methods was used in the observation of the behaviour of adaptation during captivity. In general the condition of frog captured for experiment and moved to temporary terrarium were good, in which most of them were active. All of frogs were resting before sterilization. The most effective sterilization temperature for R. margaritifer is 37˚C for 4 hours. Terrarium provided by Indonesian Safari Park were not sufficient for its movement as shown with the injured in snout part of frogs. There is only a small differences of temperature between natural habitats and terrarium which is not affecting R. margaritifer activity. Frogs were active in the evening from 18.00–23.00. During the daytime, frogs use different kind of a substrate to rest such as terrarium glass, hanjuang leafes, rocks, and fluorescent bulb. Overall, hanjuang leafes and terrarium glass was the most widely used by frogs to 23%. During 30 day period of captivity, in view from the number of feed consumption and unstable changes of body weight, frogs haven’t been able to adapt properly. Behaviour of R. margaritifer before and after sterilization suggesting that sterilization had not caused negative effect to R. margaritifer behaviour. The survival of frogs in captivity was 100% since no mortality were observed after sterilization.